BACKGROUND/AIMS: Hepatitis B and C viruses are closely associated with hepatocellular carcinoma. We studied the prevalence of infection with either virus in patients with this cancer by examination of sera and tumor tissue. METHODOLOGY: Serum samples obtained before treatment from 330 patients with hepatocellular carcinoma were assayed for antibodies against hepatitis C virus and against hepatitis B surface and core antigen. Tumor tissues from 65 patients were examined for hepatitis B virus RNA. RESULTS: Of the 330 patients, 87 had anti-hepatitis C alone; 161 had anti-hepatitis C and anti-hepatitis B (core); 13 had anti-hepatitis C and anti-hepatitis B (surface); 39 had anti-hepatitis B (surface) alone; and, 19 had anti-hepatitis B (core) alone. Eleven patients had none of these. Hepatitis B virus genes were detected in tumor tissue in all 13 patients with anti-surface antibody, in 21 of 30 patients with anti-core antibody, and in 9 of 22 patients without hepatitis B antibodies. Viral genes were detected in tumor tissue in 5 of 11 patients with neither B nor C virus markers in their sera; viral markers were found in either serum or tumor tissue in 324 of 330 patients (98.2%). CONCLUSIONS: The prevalence of hepatitis B or C virus infection in patients with hepatocellular carcinoma in Japan is extremely high. The prevalence of co-infection with both viruses is also high.